Two distinct types of HIV (human immunodeficiency virus: HIV-1 and HIV-2) have been described and are the agents responsible for AIDS. Analysis of their nucleic acid sequence has enabled different HIV-1 subtypes to be identified, although it has not been possible to establish any correlation between the variability and the pathogenicity.
The analysis of nucleotide fragments of different HIV-1 isolated has shown the existence, via analysis of the env gene, of at least 7 different subtypes designated A to G (MYERS G. et al., Human retroviruses and AIDS, 1993, Los Alamos Nat. Lab.).
More recently, another two isolates, considered to be markedly further away from the other 7 subtypes, that is to say whose sequence homology is furthest from that of the HIV-1 reference strains, have also been isolated and have been assigned to a new HIV-1 group, the O group, as opposed to the M group corresponding to the abovementioned 7 subtypes A-G, in view of their genomic organization (5' LTR Gag Pol Vif Vpu Vpr Tat Rev Env Nef LTR 3').
In view of the number of subjects suffering from HIV-1 and the mode of transmission of this virus (sexual and via blood), it was urgent to have at one's disposal both diagnostic reagents and immunogenic compositions capable of inducing the formation of neutralizing antibodies.
A number of peptides have been proposed for this purpose, originating, in particular, from the env protein or the gag protein.
As regards the gag protein, and more especially the p17 protein (included in the p55 protein), the prior art mentions a number of peptides which have been selected in the said p17 protein of HIV-1, for their diagnostic and/or vaccinal value.
There may be mentioned, for example:
International Application PCT WO 91/08227 (REPLICO AB), which relates to peptides originating from the p17 protein of HIV-1 and which essentially make it possible to discriminate between false positives and true positives. These peptides comprise the epitopes present at positions 118-132 of the p17 protein of HIV-1 (C-terminal position); PA1 U.S. Pat. No. 5,185,147 (L. D. PAPSIDERO) mentions small peptides (11 amino acids or less) which are the only ones to react with monoclonal antibodies capable of neutralizing the biological activity of HIV-1, and which are advantageous for diagnostic or vaccinal purposes (immunogenic properties). These various peptides correspond to positions 12-22 of the p17 protein; PA1 European Patent Application 0,426,314 (VIRAL TECHNOLOGIES INC and THE GEORGE WASHINGTON UNIVERSITY) mentions 5 major segments of the p17 protein of HIV-1, displaying potentially important immunogenic epitopes: peptide A (1-32), peptide B (33-50), peptide C (51-84), peptide D (85-114) and peptide E (114-131), and recommends the use of peptides A, B, C and E for the detection of HIV-1 and the use of peptides A, B, C, D and E for the preparation of an immunogenic composition. PA1 an advanced stage of the disease in the mother (grade IV), PA1 a positive P24 antigenaemia, and/or a CD4+ lymphocyte level of less than 200/mm.sup.3, PA1 a high plasma viraemia in the mother. PA1 Y5 represents AAA, AAG, GAA, PA1 Y6 represents ATA, TTA, CTG, GTA, CTA, GTG, ATG, PA1 Y7 represents GAG, GAA, PA1 Y8 represents GAA, AAG, PA1 Y9 represents ATA, GTA, CTA, GAA, GAG, GAC, GAT, AGA, PA1 Y10 represents CAA, CAG, CGA, PA1 Y11 represents AAA, AGT, AAG, AAT, CAT, AAC,ACC, PA1 Y5-Y11 have the same meaning as above, and PA1 Y1 represents AAA, AAG, PA1 Y2 represents GCT, GCC, PA1 Y3 represents TTA, CTA, PA1 Y4 represents GAT, GAC, GAG, PA1 Y12 represents AGG, AAG, AAA, ATC, AAC, PA1 Y13 represents AGC, AAC, AGT, AAG, PA1 Y14 represents GGG, AAG, AAA, CAG, PA1 Y15 represents CAA, AAA, AGA, PA1 Y16 represents AAG, AAA, ACA, CAA, AAC, AAT, PA1 Y17 represents ACA, GCA, AGC, ATA, GCC, GCT,GCG. PA1 X2 represents Lys, Glu PA1 X3 represents Ile, Leu, Val, Met PA1 X4 represents Glu, Lys PA1 X5 represents Ile, Val, Glu, Art, Leu PA1 X6 represents Gln, Arg, and PA1 X7 represents Lys, Ser, Asn, His, Thr, PA1 X2-X7 have the same meaning as above, and PA1 X1 represents Asp, Glu PA1 X8 represents Arg, Lys, Ile, Asn PA1 X9 represents Ser, Asn, Lys PA1 X10 represents Gly, Lys, Gln, Glu PA1 X11 represents Gln, Lys, Arg PA1 X12 represents Lys, Gln PA1 X13 represents Thr, Ala, Ile. PA1 Y5 represents AAA, AAG, GAA, PA1 Y6 represents ATA, TTA, CTG, GTA, CTA, GTG, ATG, PA1 Y7 represents GAG, GAA, PA1 Y8 represents GAA, AAG, PA1 Y9 represents ATA, GTA, CTA, GAA, GAG, GAG, GAT, AGA, PA1 Y10 represents CAA, CAG, CGA, PA1 Y11 represents AAA, AGT, AAG, AAT, CAT, AAC, PA1 X2 represents Lys, Glu PA1 X3 represents Ile, Leu, Val, Met PA1 X4 represents Gly, Lys PA1 X5 represents Ile, Val, Glu, Arg PA1 X6 represents Glu, Arg and PA1 X7 represents Lys, Ser, Asn, His, Thr PA1 the reagents needed for performing a PCR amplification of DNA, PA1 suitable doses of primers specific for the p17 region of the gag gene of HIV-1, and PA1 at least one nucleotide sequence according to the invention, capable of hybridizing under stringent conditions with the portion of the sequence of the p17 region of the gag gene of HIV-1 comprising one of the nucleotide sequences as defined above.
However, the collective peptides of the prior art do not solve the crucial problem of the evaluation of the risk of maternofoetal transmission of HIV-1.
Now, in 1995, the maternofoetal transmission of HIV-1 constitutes a major public health problem, since its progression goes hand in hand with that of the contamination with HIV-1 throughout the world.
The World Health Organization (WEO) estimates at more than one million the number of children contaminated worldwide.
In France, the number of women of child-bearing age who are seropositive for HIV-1 is estimated at 30,000-40,000.
The degree of vertical transmission is from 15 to 25% in both Europe and the USA. This degree is higher on the African continent, ranging from 20 to 40% for reasons which are not very clear, doubtless as a result of coinfection of the mother and breast-feeding.
Putting European and American pregnant women on Retrovir.sup.(R) (AZT) until they give birth has enabled the degree of transmission to be reduced to 8.3%.
Twelve years after the discovery of the virus, our ignorance of the physiopathology of the vertical transmission is still considerable: many arguments indicate that transmission of the virus from mother to foetus may take place at a late stage in utero (in the last two months), at the time of delivery with a probably greater incidence and during breast-feeding, which is consequently strongly discouraged.
Among the parameters which may influence the vertical transmission, few have, at the present time, been demonstrated as being positively correlated with the risk of transmission. These are:
The absence of one of these parameters does not, however, rule out the risk of vertical transmission, it merely decreases it.
In contrast, the presence of neutralizing antibodies in the mother, protecting passage to the child, remains very controversial.
Among the virological factors of biological expression (degree of replication, cellular tropism, cytotoxicity, capacity for induction of syncytia) or molecular factors of biological expression (in particular in the gene coding for the gp120 envelope glycoprotein), none has, to date, been demonstrated as being able to influence the risk of vertical transmission (P. ROQUES et al., AIDS, 1993, 7 (suppl. 2), S39-S43; F. X. MBOPI KEOU et al., Bull. Inst. Pasteur, 1994, 92, 3-18; Y. F. KHOURI et al., J. Clin. Invest., 1995, 95, 732-737; G. A. MULDER-KAMPINGA et al., Journal of Virology, 1995, 69, 2285-2296).
It was consequently the object of the present invention to provide for nucleic acid fragments and the corresponding peptides capable of serving as reagents for detection and evaluation of the risk of maternofoetal transmission of HIV-1.